Hepatitis a virus (HAV) is a picornavirus with a single-stranded RNA genome of approximately 7500 nucleotides. The wild-type strain of HAV grows poorly in cell-culture, is not cytopathic, and yields a low titer of virus. A cell-culture adapted mutant has been isolated which grows significantly more efficiently in cell-culture and which is attenuated for marmosets and chimpanzees. The objectives of this project are to determine the genetic basis for virulence and adaptation to cell culture of HAV. 1. The molecular basis of the ability of attenuated HAV strain, HM-175, to grow in cell culture has been determined. Growth in cell culture is dependent upon three mutations in the 2B and 2C regions of the HAV genome. Host cell specificity is conferred by changes in the 5' noncoding region. 2. A deletion in the non-infectious cDNA clone containing the genome of wild-type virus was identified and corrected and the new cDNA clone was shown to be infectious for cultured cells and to produce virus that caused severe hepatitis in marmosets. 3. Simian strain AGM-27 was recovered from an infected, ill African green monkey. This HAV strain was evaluated for host range and virulence in several primate species. It is attenuated for chimpanzees but not African green monkeys or marmosets and is being further evaluated as a candidate live hepatitis A vaccine. It has been almost entirely sequenced and found to be significantly different from human HAV strains.